Channelopathies, genetic disorders in brain and muscle ion channels, are useful to investigate the structural determinants of channel function. In the present proposal we will use point mutations of neuronal and skeletal muscle sodium channels to determine the mechanisms for channel functions not well understood, including deactivation and closed-state fast inactivation. We will focus on identifying allosteric, domain-specific regulation of S4 voltage sensors in Specific Aims I to III. We will determine the domain-specific involvement of the voltage sensors in a novel form of channel closure, and then of cytoplasmic regions in the poorly understood process of closed-state fast inactivation. We will take advantage of the fact that these channelopathies provide point mutations in multiple channel domains. These data will support the long-term objectives of the laboratory of the PI to study novel channel functions, and novel mechanisms of channel dysfunction in ion channel diseases such as myotonia and epilepsy. Undergraduates will be trained in molecular biology and patch clamp electrophysiology, and asked to design progressive, testable hypotheses on deactivation and closed-state fast inactivation. A graduate student will construct and test the effects of C terminal mutations on the kinetics and completion of closed-state fast inactivation. Together with the PI, that student will then use gating current measurements to determine the mechanisms by which the C terminus and S4-S5 linkers regulate DIII and DIV voltage sensor movements in closed-state fast inactivation. PUBLIC HEALTH RELEVANCE: Genetic disorders consisting of specific mutations in ion channel genes are called "channelopathies". The defects in ion channel function in these disorders can be used to illuminate the structural determinants within the sodium channel that allow this protein to control electrical signaling in the brain and periphery. These studies also provide insight into the molecular basis of genetic disorders.